This Phase II STTR application is a continuation of a previous Phase I award between the Wright group at Dartmouth College and Promiliad Biopharma. The goal of this application is to continue our efforts to develop novel chemical libraries with the diversity and complexity common to natural products as a marketable product to the pharmaceutical industry. Key to our strategy is the use of fermentation derived natural products and recombinant non-natural variants as high complexity feedstock materials for diversity oriented synthesis. Unlike analog generation, these recombinant natural products are specifically designed to allow chemical diversification. Our approach differs substantially from combinatorial biosynthesis in that only a few, well-defined compounds are needed from fermentation sources. By only requiring a limited set of recombinant compounds from the organism, careful optimization can be conducted to ensure high yields of material. While this strategy relies on nature to construct a high degree of the complex features of the library, diversity-oriented strategies are utilized to generate large libraries that cover vast areas of structural space. Under the Phase I award, we have developed a number of direct and efficient strategies for converting a recombinant building block (nonactate) into several unique molecular architectures. This Phase award will allow our efforts to transition from the methods stage to Phase III, large-scale production and marketing of compound libraries. Under this award we will (1) prepare moderate sized libraries of highly diverse and complex molecules from nonactate, (2) develop new methods for scaffolds generation from the polyketides methymycin, pikromycin and spinosyn and (3) develop two new recombinant natural feedstocks, pikrodiene and spinosene. Through these efforts, Promiliad will be in a position to become a major supplier of new chemical entities to the pharmaceutical industry. [unreadable] [unreadable] [unreadable]